Abstract In Vitro Diagnostic Solutions (IVDS) proposes to develop the first and only FDA-approved point-of-care test (POCT) for the determination of L-phenylalanine (Phe) from a finger or heel-stick sample. The PKU Now will serve as a lifelong home monitoring device for PKU patients as well as a diagnostic testing device in lesser- developed regions. It will provide immediate feedback on blood Phe levels, which will enable stricter dietary compliance, substantially improve the quality of life for PKU patients and reduce healthcare costs. In Phase I studies, we successfully resolved three critical issues. We eliminated tyrosine interference that could obscure the measurement of Phe (Aim-1), eliminated hematocrit bias in the range of 32% to 60% (Aim-2) and established concordance between the ?PKU Now? and a laboratory reference method, Amino Acid Analysis (AAA), in terms of accuracy and precision (Aim-3). PKU is the most common amino acid disorder, affecting 1 in 10,000 people. Most cases of PKU are detected by newborn screening in developed countries. PKU patients must monitor and control their Phe levels throughout their lives to avoid severe neurological complications, permanent intellectual disabilities and delayed development. Currently, Phe levels are measured at laboratories via blood collection or blood spotted onto Dried Blood Spot Cards and sent for mass spec analysis. This complicated process for monitoring Phe makes real time measurements impossible resulting in non-compliance. ?Currently, results from DBS testing can take days to weeks. This situation endangers the health and well-being of tens of thousands of people with PKU. Such a device will open the way for newborn screening of PKU in developing countries. Of all babies born around the world, more than 2/3 won?t be tested for PKU? (PKU.org). IVDS?s proposed POCT, the ?PKU Now?, will provide real-time blood Phe levels, thereby enabling immediate intervention and improving the quality of life of thousands of people. To complete development of the PKU Now, we will: Aim-1: Finalize assay development by evaluating bioactive components; Aim-2: Transition from hand assembly to semi-automated assembly of test strips; Aim-3: Develop and test synthetic controls; Aim-4: Validate the PKU Now using spiked blood samples (following potential changes from Aim-1); and Aim-5: In two Blind Studies, we will establish concordance between the data obtained from the PKU NowTM and data obtained from AAA using PKU patient samples. These studies will be conducted under approved IRBs? and will follow all HIPPA guidelines. Once commercialized, the PKU NowTM will be the first POC diagnostic tool and home monitoring device for PKU patients, enabling real-time intervention and treatment.